Chromatography Flashcards
How do all types of chromatography work?
There is a mobile phase
There is a stationary phase
Separation depends on the attraction to each phase and each component will have a different retention time / Rf value
Why does separation occur
Polarity
One phase will be polar and one will be non polar
Polar substances dissolve well in polar solvents and vice verse
What is the method for thin layer chromatography
1) draw line 1.5cm from bottom of TCL plate
2) Place a drop of purified solid on watch glass and dissolve on a few drops of solvent
3) use capillary tube to place spot on pencil cross
4) allow spot to dry and repeat for solid
5) Place solvent phase in beaker to depth of 1cm
6) place TLC plate in beaker and cover with lid
7) allow solvent to rise and mark solvent front
8) place in fume cupboard until it is dry and solvent has evaporated
9) shine UV light
10) calculate Rf value
What width must the spot be in chromatography
No larger than 0.5 cm
To produce a concentrated spot
How to calculate Rf value
Distance moved by spot
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Distance moved by solvent front
Other than UV light how else can you view colourless spots after chromatography
Spray with developing agent eg ninhydrin
Place plate in a beaker containing iodine crystals and cover with a watch glass
Iodine is a locating agent which caused the spots to become brown
Why is it necessary to wear plastic gloves when holding a TLC plate
To avoid contamination from amino acids on the skin
Why a pencil line?
Insoluble so will not move with solvent
How is a very tiny concentrated drop of amino acid added to the TLC plate?
With a capillary tube
Allow it to dry and repeat
Why does the beaker solvent have a depth of only 1cm
If it is too deep it will dissolve the mixture
Why is the beaker sealed with a lid when the TLC plate is placed on it
Solvent may evaporate
A sealed environment allows the solvent to saturate the atmosphere inside
Why is the TLC plate left to dry in a time cupboard?
Solvent may be flammable or toxic
What is ninhydrin?
What are the hazards of it and how are they avoided?
A developing agent
Skin irritant
Wear gloves, use fume cupboard, avoid breathing in the vapour
